PROJECT SUMMARY In recent years, there has been considerable interest in the possible role that members of the transforming growth factor- family may play in regulating tissue aging and the possibility that manipulating their levels of signaling may be a new therapeutic strategy to combat tissue dysfunction in the elderly. Much of this interest has focused on two highly related signaling molecules, myostatin (MSTN, GDF-8) and GDF-11, both of which were originally identified by my laboratory many years ago. We showed that MSTN normally acts to limit muscle growth, and as a result, there has been considerable interest in the possibility that inhibitors of MSTN signaling might be effective in enhancing muscle strength and regeneration; in this respect, there are currently at least 11 phase II or phase III clinical trials being conducted by 7 pharmaceutical and biotechnology companies testing MSTN inhibitors in patients with muscle loss, including in patients who have undergone hip replacement surgery resulting from falls as well as in the elderly with age-related sarcopenia. In the case of GDF-11, recent studies by other groups have suggested that GDF-11 may play an important role in tissue aging. Specifically, several papers reported that circulating GDF-11 levels decrease as a function of age and that systemic administration of purified GDF-11 protein can reverse age-related tissue dysfunction in the heart, skeletal muscle, and nervous system. Other studies, however, reported the opposite, namely, that GDF- 11 circulating levels do not decrease with aging and that administering GDF-11 protein has a detrimental effect on muscle regeneration. Clearly, elucidating the roles of these signaling molecules in regulating adult tissue homeostasis will be critical not only to understanding the control of tissue aging but also to the development of therapeutic strategies for manipulating the activities of these molecules for clinical applications in the elderly. In this project, we will attempt to elucidate the roles of this signaling pathway in aging skeletal muscle by focusing on MSTN, GDF-11, and the related ligand, activin A as well as their inhibitory binding proteins, follistatin (FST), FSTL-3, GASP-1, and GASP-2, all of which circulate in the blood. The overall goal of this project is to determine whether these ligands and binding proteins are pro-geronic or anti-geronic. For this project, we will take advantage of the many genetic and pharmacological tools that we have developed over many years targeting the various components of this regulatory network. The Specific Aims are: to determine how circulating levels of MSTN, GDF-11, and activin A and their inhibitory binding proteins change as a function of age in mice and how their expression patterns in skeletal muscle following injury differ in aged versus young mice; to use mouse lines carrying targeted mutations in genes encoding these ligands and their binding proteins to examine the roles of these proteins in regulating skeletal muscle and other tissues in aged mice; and to use genetic and pharmacological approaches in conjunction with parabiosis studies to examine effects of targeting these ligands and binding proteins on skeletal muscle regeneration in aged mice.